The availability of cytosolic ATP is essential for myocardial contraction and relaxation, and the depletion of ATP leads to irreversible ischemic damage during reperfusion after an ischemic insult. Thus, the clinical management of myocardial ischemia and cardiac protection during coronary bypass should be aimed at preventing depletion of ATP. In the heart, an imbalance between oxygen supply and demand causes an immediate fall in PCr, followed by a net hydrolysis of ATP and an increase in the concentration of AMP which is then dephosphorylated to adenosine by the enzyme 5'nucleotidase, or deaminated to IMP by the enzyme AMP deaminase. Adenosine is membrane permeable and the release of adenosine evokes many cardioprotective mechanisms. Furthermore, AMP hydrolysis to adenosine provides a mechanism whereby the phosphorylation potential is preserved by mass balance during compromised energy supply (i.e. low energy nucleotides are removed). Clearly, AMP hydrolysis is beneficial during acute ischemia, yet the continuous loss of nucleosides during prolonged ischemia will lead to the depletion of nucleotide pools. The regulation of AMP hydrolysis is therefore important for myocardial survival during prolonged ischemia and cardioplegia. We have previously shown that AMP hydrolysis is downregulated during prolonged underperfusion. The experimental data showed depressed purine efflux at similar cytosolic AMP concentrations during the second of two identical periods of underperfusion and analysis with a mathematical model indicated downregulation of 5'nucleotidase (5NT) late in the first period of underperfusion. Since adenosine release aids in the adaptation to acute hypoxic stress, we have addressed the question whether the downregulation of 5NT persists during an increase in the substrate AMP, evoked by a period of greater hypoxic stress. In order to test the hypothesis that 5NT is downregulated during prolonged underperfusion yet becomes upregulated, or un-downregulated, during greater hypoxic stress, experiments were designed in which cytosolic AMP levels were greatly increased by applying hypoxia. Evidence has also been accumulating in the literature that the rate of adenosine formation in the cytosol is not solely dependent on 5NT activity, but is the result of at least four factors: the AMP concentration, and the activities of three enzymes which influence this concentration: adenosine kinase (AK), which rephosphorylates adenosine to AMP, AMP deaminase, which deaminates AMP to IMP, and finally AMP-preferring 5'nucleotidase, which dephosphorylates AMP to adenosine. Since our previous mathematical model did not describe AMP deaminase, this pathway, along with the IMP-preferring isoform of 5NT, has been included in the present model.